The lysosome is an organelle founded in the cytoplasm of eukaryotic cells, which serves as storage for many hydrolytic enzymes and as a center for degrading and recycling cellular components. This organelle contains several types of hydrolytic enzymes, including proteases, nucleases, glycosidases, lipases, phospholipases, phosphatases and sulfatases. All enzymes are acid hydrolases.
Lysosomal storage diseases (LSDs) are caused by genetic defects that affect one or more lysosomal enzymes. These genetic diseases result generally from a deficiency in a particular enzyme activity present in the lysosome. To a lesser extent, these diseases may be due to deficiencies in proteins involved in lysosomal biogenesis.
LSDs are individually rare, although as a group these disorders are relatively common in the general population. The combined prevalence of LSDs is approximately 1 per 5,000 live births. See Meikle P, et al., JAMA 1999; 281:249-254. However, some groups within the general population are particularly afflicted by a high occurrence of LSDs. For instance, the prevalence rates of the Gaucher and Tay-Sachs diseases in descendants from Jewish Central and Eastern European (Ashkenazi) individuals is 1 per 600 and 1 per 3,900 births, respectively. The Finnish population is also afflicted by an uncommonly high LSDs prevalence rate.
Type III mucopolysaccharidoses (MPSIII), known collectively as Sanfilippo syndrome, are LSDs caused by a deficiency in one of the enzymes involved in the degradation of heparan sulfate, leading to its pathological accumulation. MPSIII is classified into four subtypes depending on the enzyme deficiency. Loss of sulfamidase activity causes subtype IIIA and has been reported to be the most severe, with the earliest disease onset and shortest survival. Symptoms of MPSIIIA occur in the first years of life, and are characterized by severe neurodegeneration that leads to deep mental retardation, aggressiveness, hyperactivity, and sleep alterations. Patients progressively lose the capacity of speech, swallow, and basic motor coordination. In addition to the neurological symptoms, MPSIIIA patients suffer non-neurological alterations, including hepato- and splenomegaly, skeletal and joint malformations, as well as frequent diarrhoea and respiratory tract infections. The progressive worsening of symptoms results in the death of the patient during adolescence. See Neufeld E, Muenzer J, “The mucopolysaccharidoses” in Scriver C, et al., Eds., “The metabolic and molecular basis of inherited disease” (McGraw-Hill Publishing Co., New York, N.Y., US, 2001, pp. 3421-3452).
There is no cure for MPSIIIA currently and, therefore, existing treatments are aimed at controlling symptoms of the disease in order to improve the poor quality of life of the patients. MPS disorders can be treated by bone marrow transplantation or enzyme replacement therapy (ERT). Both approaches rely in the endocytosis of lysosomal enzymes from extracellular medium and their targeting to lysosomes via the mannose-6-phosphate receptor (M6PR) present at the cell membrane. Nevertheless, bone marrow transplantation has demonstrated to be inefficient in the treatment of MPSIII patients. See Sivakamur P, Wraith J, J. Inherit. Metab. Dis. 1999; 22:849-850. ERT has been extensively proven to be effective in counteracting the non-neurological accumulation in other lysosomal storage diseases, including MPSI, II and VI. See Harmatz P, et al., J. Mol. Genet. Metab. 2008; 94:469-475; Muenzer J, et al., Genet. Med. 2006; 8:465-473 and Wraith J, et al., J. Pediatr. 2004; 144:581-588. In addition to the high cost of these treatments, it has been shown that ERT does not result in correction or preservation of neuronal function due to the insufficient delivery of the exogenously provided enzyme through the blood-brain barrier (BBB). See Enns G, Huhn S, Neurosurg. Focus 2008; 24:E12. More recently, it has been demonstrated that high-dose ERT is partially successful in clearing CNS storage in MPS VII, possibly due to the saturation of M6PR and mannose receptors that lead to a longer half-life of the protein in circulation. See Vogler C, et al., Proc. Natl. Acad. Sci. USA 2005; 102:14777-14782. This study demonstrates that high levels of the enzyme in circulation during long periods of time correlate with a better correction of the pathology. Intracerebral and intra-CSF delivery of the enzyme have also been proved to be efficient in reducing CNS pathology in MPS IIIA mice. See Hemsley K, et al., Genes Brain Behav. 2008; 53(2):161-8 and Saves P, et al., Mol. Genet. Metab. 2004; 82:273-285. However, this approach is highly invasive due to the need for multiple repeated injections and could increase the risk of damage and/or infections in the brain.
Given the limitations of current therapeutic options for MPSIII, alternative approaches are needed. Gene transfer could provide the means to achieve a permanent production of the missing enzyme from a single intervention. Adeno-associated vectors (AAV) are rapidly emerging as the vector of choice for many gene therapy applications, due to their high transduction efficiency and their lack of pathogenicity. AAV vectors efficiently transduce post-mitotic cells and several pre-clinical and clinical studies demonstrated the potential of AAV vector-mediated gene transfer to efficiently drive sustained expression of therapeutic transgenes for a variety of diseases. See Daya S, Berns K, Clin. Microbiol. Rev. 2008; 21:583-593.
It has been shown that the administration of an AAV5 vector co-expressing sulfamidase and the sulfatase activator SUMF1 in lateral ventricles of newborn MPSIIIA mice is able to correct many neurological and behavioral alterations. See Fraldi A, et al., Hum. Mol. Genet. 2007; 16:2693-2702. However, this proposed course of action has several shortcomings. First, the CMV promoter utilized has been reported to silence. Second, the long term effects of the co-expression of sulfamidase and SUMF1 have not been assessed yet. It is not clear if the co-expression of SUMF is even necessary and provides any additional permanent benefits in comparison to the treatment with sulfamidase only. Third, AAV5 vectors have a low distribution within the parenchyma, and more importantly, the delivery of sulfamidase into the brain by using these vectors does not result in any transduction of the cerebral tissue, thus, no correction of somatic phenotype is achieved by following this approach. Finally, Fraldi, 2007, supra demonstrated the efficacy of gene transfer in only newborn MPSIIIA mice. No experiments were reported in older mice. Since MPSIIIA is usually diagnosed after 3-4 years of age, the newborn animal model is not adequate for predicting the effects of this treatment in human beings.
In view of the difficulties for diagnosing MPSIIIA at birth, the development of therapeutic interventions starting in early adulthood has been proposed. It has been reported that the intravenous delivery of a lentiviral vector expressing sulfamidase in adult mice resulted in little amelioration of the CNS phenotype, likely due to the relatively poor transduction performance of these vectors in vivo. See McIntyre C, et al., Mol. Genet. Metab. 2008; 93:411-418. Thus, the use of viral vectors with higher transduction efficacy in vivo, such as AAV vectors, may provide higher circulating levels of sulfamidase, which could potentially ameliorate or correct the neurological pathology.
The treatment of MPSIIIA via gene therapy requires more efficient vectors and sulfamidase coding sequences. Therefore, there is a long-felt need for an effective treatment of MPSIIIA. There is also the need for novel approaches to the treatment of this disease that would have enhanced security features. MPSIIIA is a rare disease and is therefore an orphan disease. The pharmaceutical agents developed specifically to treat this rare medical condition will be orphan drugs.